It is proposed by E. J. Cohn in U.S. Pat. No. 2,390,074 that proteins useful in blood transfusion technology be produced by a blood plasma fractionation process in which an organic solvent such as alcohol is employed as a precipitant. The method proposed by Cohn depends on balancing the precipitating action of the organic solvent with the solvent actions of the electrolytes present, whereby a series of conditions may be established at which the solubility of any particular protein will remain relatively constant. The solubility of certain other proteins may in some of these conditions be such that reasonably pure separations may be possible.
In such a method five independent variables are usually controlled:
1. electrolyte concentration PA1 2. alcohol concentration PA1 B 3. hydrogen ion concentration PA1 4. temperature PA1 5. protein concentration
Cohn teaches that when a readily denatured protein e.g. a globulin is to be precipitated, considerable care should be exercised in the addition of the precipitant to the plasma or subfraction thereof. Thus it is recommended that after suitable pH and temperature adjustment of the plasma (or subfraction thereof) the precipitant be added thereto by way of a semi-permeable membrane to avoid denaturation. It will be appreciated that the process is essentially slow and the concentration of precipitant varies continuously up to the point at which all the precipitant has been added. In consequence precipitation of fractions takes place en route and a long period of ageing is required in order to approach final equilibrium. It is in practice very rare to achieve the equilibrium condition required and the final product is almost invariably contaminated. Such bulk systems also have the disadvantage that large volumes of plasma liquor are continually at risk of plant failure or staff errors. A large ageing period has moreover been considered necessary in the past in order to achieve protein fractions having a physical form which permits ease of recovery by centrifugation, filtration or other standard methods.
Cohn also teaches that where the desired product is a less labile protein such as albumin, alternative procedures may be employed for the addition of precipitant. Plasma and precipitant may be for example supplied separately to each end of a T-shaped tube, mixing at the junction and during passage down the stem. In such a system there is however a substantial risk of obstruction by precipitated protein and the process may thereby be rendered inoperative.